Atherosclerosis is an excessive inflammatory/proliferative response of the vascular wall to various forms of injury. It has been suggested that, during inflammation, reactive oxygen (ROS) and reactive nitrogen species (RNS)-induced endothelial cell damage represent an important primary event in the process of atherosclerotic lesion formation. The resulting oxidative and nitrosative stress impairs the critical balance of the availability of endothelium-derived nitric oxide in turn promoting the proinflammatory signaling events, ultimately leading to the plaque formation. Atherosclerosis initiating events may be different under different conditions; however endothelial dysfunction is known to be one of the major initiating events. Macrophages also undergo apoptosis inside the endothelium, leading to their phagocytic clearance.
Increased mitochondrial oxidative damage is a major feature of most age-related human diseases including atherosclerosis and abnormal electron leakage from mitochondria in the respiratory chain in oxidant-stressed cells triggers the formation of ROS in mitochondria leading to altered behavior of the cell/cell death. Previously many studies have linked excess generation of ROS with vascular lesion formation and functional defects. More so, a role for mitochondria-derived ROS in atherogenesis is supported by links between common risk factors for coronary artery disease and increased levels of ROS. Mitochondrial ROS is increased in response to many atherosclerosis inducers including hyperglycemia, triglycerides and ox-LDL. Aortic samples from atherosclerotic patients had greater mitochondrial DNA (mtDNA) damage than nonatherosclerotic aortic samples from age-matched transplant donors (Mitochondrial integrity and function in atherogenesis. Circulation. 2002; 106:544-549). Even though endothelial cells have low mitochondria content, mitochondrial dynamics acts as a prime orchestrator of endothelial homeostasis under normal conditions, an impairment of mitochondrial dynamics because of excess ROS production would cause endothelial dysfunction resulting in diverse vascular diseases. Exposure of endothelial cells to free fatty acids, a common feature seen in patients with metabolic syndrome increases mitochondrial ROS (Palmitate induces C-reactive protein expression in human aortic endothelial cells. Relevance to fatty acid-induced endothelial dysfunction. Metabolism. (2011) 60: 640-648).
Therefore keeping in view of the involvement of mitochondrial ROS in causing endothelial dysfunction leading to the accentuation of vascular diseases, it would be ideal to counteract mitochondria ROS by targeting ROS scavengers specifically to the site of action. The major drawback of antioxidant therapy in the treatment of mitochondrial diseases has been the inability to enhance antioxidant levels in mitochondria. Recently, there was a breakthrough in mitochondrial targeting of antioxidants (Drug delivery to mitochondria: the key to mitochondrial medicine. Adv Drug Deliv Rev. (2000) 41: 235-50). Antioxidants were covalently coupled to a triphenylphosphonium cation (TPP+), and these compounds were preferentially taken up by mitochondria (Selective targeting of a redox-active ubiquinone to mitochondria within cells: antioxidant and antiapoptotic properties. J Biol Chem. (2001) 276: 4588-4596). The lipophilic cations easily permeate through the lipid bilayers and subsequently build up several hundred-fold within mitochondria because of a large mitochondrial membrane potential. The uptake of lipophilic cations into the mitochondria increases 10-fold for every 61.5 mV difference in the membrane potential, leading to a 100- to 500-fold accumulation in mitochondria. This strategy not only reduces the concentration of the molecule that is being employed to scavenge ROS, but also reduces the non-specific effects of the molecule if it were to be used at high concentrations to elicit a similar effect.
Coumarins constitute a group of phenolic compounds widely distributed in natural products (The Pharmacology, Metabolism, Analysis and Applications of Coumarin and Coumarin-Related Compounds. Drug Metab Rev (1990) 22: 503-529), and they have recently attracted much attention because of their wider pharmacological activities. Of these, esculetin (6,7-dihydroxycoumarin) has been shown to be a lipoxygenase inhibitor, and it inhibits the production of leukotrienes and hydroxyeicosatetraenoic acid through the lipoxygenase pathway. More recently, esculetin has been reported to inhibit oxidative damage induced by tert-butyl hydroperoxide in rat liver (Inhibitory effect of esculetin on oxidative damage induced by t-butyl hydroperoxide in rat liver. Arch Toxicol. (2000) 74:467-72). Esculetin protects against cytotoxicity induced by linoleic acid hydroperoxide in HUVEC cells and the radical scavenging ability of esculetin was confirmed by electron para magnetic resonance spectroscopy (Protection of coumarins against linoleic acid hydroperoxide-induced cytotoxicity. Chemico-Biological Interactions 142 (2003) 239-254). However, as coumarins may have poor bioavailability in vivo and do not significantly accumulate within mitochondria, their effectiveness remains limited and because of this, they may have to be employed in higher concentrations to scavenge mitochondrial ROS. In the present patent application, we have used lipophilic cation (TPP+) to target esculetin (FIG. X) to mitochondria and show that mitochondria-targeted esculetin (Mito-Esc) protects oxidant-induced endothelial cell death via nitric oxide and AMPK-dependent pathways at far below concentrations than reported earlier with native esculetin and further we report that Mito-Esc significantly inhibits aortic aneurysm (AA) and atheromatous plaque formation in Angiotensin-II-induced atherosclerotic process in Apolipoprotein E−/− mice model. The following are the prior art literature related to the present invention (WO1996031206; U.S. Pat. No. 6,331,532; WO2008145116; U.S. Pat. No. 4,977,276; U.S. Pat. No. 4,230,624; WO2011115819).